Genetic basis of chloroquine resistance. Linkage analysis indicates that chloroquine resistance in P. falciparum is controlled by a single gene or closely linked group of genes within a (equals approximately) 200 kilobase segment on the parasite's seventh chromosome. We are using yeast artificial chromosomes to map this segment in detail and are identifying transcribed segments that should lead to a candidate resistance gene. Other drug development targets. Studies have progressed on two Plasmodium enzymes: dihydrofolate reductase-thymidylate synthase (DHFR-TS) and S-adenosylhomocysteine hydrolase (AHCY). Knowledge of point mutations in DHFR-TS is being used as a basis for epidemiological surveys and for development of alternative drugs. The predicted amino acid sequence of P. falciparum AHCY is now determined. Heterologous expression and purification of the enzyme is being pursued for kinetic and functional comparisons with mammalian AHCY. Transfection of Plasmodium. Efforts are focused on development of constructs and on DNA delivery methods for parasite transfection. Stable transfection is being attempted with a dihydrofolate reductase-thymidylate synthase (DHFR-TS) construct. Transient expression experiments are being pursued with luciferase constructs. Ballistic particles and electroporation are being evaluated for DNA delivery. Genetic analysis of red cell invasion. A switch to an alternative pathway of red cell invasion was detected in a P. falciparum clone made to propagate in neuraminidase-treated, sialic acid-deficient red blood cells. The switch may involve a mechanism in which certain genes are activated or rearranged. Differential and subtractive screening methods have so far been unsuccessful. An alternative approach using genetic linkage analysis is being examined.